The grant, which was presented by the Academic Enrichment Funds from the Dean’s Office at the CU School of Medicine, will optimize the pipeline to build custom Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) libraries for labs at the CU Cancer Center and all over the state. These CRISPR libraries help researchers edit genes in cancer cells to learn more about how these genes work with or against cancer.

“We are so excited to receive this grant,” says Molishree Joshi, PhD, manager and consulting scientist for the FGSR. “We were already planning on building custom pools and this grant motivates us even more to get it done.”

Background

The goal of functional genomics and the FGSR is to help understand the roles of specific genes. Because the genes found in your DNA are the blueprints for proteins, turning them up or down, on or off, can result in more, less or different proteins building cells and controlling their behavior. This is important when it comes to cancer research because if a gene becomes mutated it can create abnormal proteins that lead to cancerous cells. By enhancing or suppressing a gene, researchers in the FGSR hope to understand what affects it could have on that cell.

What is CRISPR?

One of the latest technologies that allows scientist to enhance or suppress a gene is by using the CRISPR-Cas9 system.

The CRISPR-Cas9 system uses two tools to allow researchers to snip a specific part of DNA sequences in a huge genome — the Cas9 protein and a guide RNA (gRNA). The gRNA matches the sequences of a target gene, directing the Cas9 protein to a specific part of the DNA where Cas9 makes a cut. The cell’s native DNA repair machinery generally repairs the cut — but often makes mistakes, thereby editing the specific gene and often times making it non-functional. Although the FGSR has more than 35,000 individual ready-to-use gRNAs to guide gene editing of every gene in the human genome, and multiple pooled genome-wide CRISPR libraries, the ability to create custom CRISPR pools will allow the team at the FGSR, and hence the researchers at University of Colorado, to target a specific set of genes and evaluate them in a complex genetic screen. A CRISPR pool is collection gRNAs designed and tested together in a cell line or model organism to identify the gene(s) essential for a given phenotype. By conducting experiments with these different pools, researchers can learn why cells become therapy resistant and identify future targeted therapies.

“We have had many inquiries about customized pools but have lacked the setup,” says Joshi. “Because of this grant, we are very encouraged to begin streamlining the process to deliver sequence-verified custom CRISPR pools, locally and affordably.”

Custom CRISPR pools will allow researchers to validate genome-wide screens by targeting a smaller number of genes. Additionally, researchers that focus on a specific pathway will have the ability to perform smaller focused screens using a greater number of gRNA targeting each gene for higher coverage.

Joshi and her team have already started the computational work to create customizable pools and hope to have the first library done soon.

“We are hoping to have the first library ready for testing by March,” she says. “Once that is done we can start creating libraries for other labs to test.”

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